Nitrogen fixation and nifH diversity in human gut microbiota.

Katsura Igai,Ayako Morita,Suguru Nishijima,Paul F Horwood,Jun Baba,Kiyoshi Tadokoro,Kiwamu Minamisawa,Kazumi Natsuhara,Shingo Odani,Taro Yamamoto,Andrew R Greenhill,Eriko Tomitsuka,Masahira Hattori,Takumi Tsutaya,Masahiro Umezaki,Moriya Ohkuma,Yuichi Hongoh,Hirohito Tsurumaru,Wataru Suda,Manabu Itakura,Minoru Yoneda,Jun Inoue ,Peter Siba

doi:10.1038/srep31942

Abstract

It has been hypothesized that nitrogen fixation occurs in the human gut. However, whether the gut microbiota truly has this potential remains unclear. We investigated the nitrogen-fixing activity and diversity of the nitrogenase reductase (NifH) genes in the faecal microbiota of humans, focusing on Papua New Guinean and Japanese individuals with low to high habitual nitrogen intake. A 15N2 incorporation assay showed significant enrichment of 15N in all faecal samples, irrespective of the host nitrogen intake, which was also supported by an acetylene reduction assay. The fixed nitrogen corresponded to 0.01% of the standard nitrogen requirement for humans, although our data implied that the contribution in the gut in vivo might be higher than this value. The nifH genes recovered in cloning and metagenomic analyses were classified in two clusters: one comprising sequences almost identical to Klebsiella sequences and the other related to sequences of Clostridiales members. These results are consistent with an analysis of databases of faecal metagenomes from other human populations. Collectively, the human gut microbiota has a potential for nitrogen fixation, which may be attributable to Klebsiella and Clostridiales strains, although no evidence was found that the nitrogen-fixing activity substantially contributes to the host nitrogen balance.

Highlights

Rumen contents or faeces of herbivorous mammals, such as sheep[8,9,10], goats[8,10], cows[11,12], rabbits[8], and reindeer[8]
We have demonstrated that the human gut microbiota truly has a capacity for nitrogen fixation, using 15N2 incorporation and acetylene reduction assays
The acetylene-reduction rates in the human faecal samples were up to 1.6 nmol/g/h, which is comparable to those detected in herbivorous mammals, such as sheep (0–1.4 nmol/g/h in the rumen)[8,9,10], a rabbit (0.38 nmol/g/h in the cecum)[8], and a reindeer (0–0.77 nmol/g/h in faeces)[8]

Summary

Introduction

Rumen contents or faeces of herbivorous mammals, such as sheep[8,9,10], goats[8,10], cows[11,12], rabbits[8], and reindeer[8]. In a study of people with a low-nitrogen diet in Papua New Guinea (PNG), Bergersen and Hipsley (1970) hypothesized that nitrogen fixation occurs in the human distal gut[14]. The habitual nitrogen intake of the study subjects was estimated to be 49–74 mg/kg body weight/day in 196716, which is much lower than the estimated average requirement for humans (105 mg/kg body weight/day)[17] These individuals displayed no disorders associated with nitrogen (protein) deficiency[18]. Bergersen and Hipsley collected faeces from an unspecified number of PNG individuals and five Europeans, and analysed them with an acetylene reduction assay. They detected 0.75 nmol/g/h acetylene-reducing activity in one of the European faecal samples and ‘slight but significant’ activities in all the PNG samples. This study demonstrates the nitrogen-fixing activity of the human faecal microbiota and presents a list of candidate nitrogen-fixing bacteria in the human gut

Methods

Results

Conclusion